A portion of the East Village of Angel Mounds near Evansville, Ind., under excavation in 1941 by Works Progress Administration workers led by Indiana archaeologist Glenn A. Black. Black proposed that a pair of co-buried infant skeletons excavated at the time were conjoined twins. Image courtesy Indiana University.

Were they, or weren’t they?

The question had plagued archaeologists at Indiana University’s Glenn Black Laboratory for decades. Ever since Glenn Black himself dug up the weird and unique burial site back in 1941, the question has only grown, even into mythical proportions.

The burial site in question was of a pair of infants at the Middle Mississippian Age (A.D. 1050-1400) village near Evansville, Indiana, which is near the southwestern tip of the state on the Ohio River. Many societies at the time viewed twin births negatively and one or both twins would be killed, while in other Eastern North American societies, a twin birth was accorded high status and deaths would have warranted excessive ceremony.

So the fact that two infants were in the same burial spot and yet was unremarkable otherwise with no adornment and a location in the common burial area at Angel Mounds led Black to one conclusion – the two must have been conjoined twins. Even though there were no signs of skeletal fusion, he reasoned they still could have been flesh-joined twins.

But of course, nobody could ever know. At least, that is, until modern DNA sequencing techniques progressed to the point where some tests could be run.

The only DNA that reasonably could be extracted, however, was maternally inherited mitochondrial DNA, or mtDNA. If you remember back to middle school, mitochondria are the tiny organelles within our cells that are “cellular power plants” in that they generate adenosine triphosphate (ATP), used as a source of chemical energy.

Billions of years ago, when all life consisted of single microscopic cells, the theory goes that one of these cells swallowed another. Typically, this would spell the end of the line for the engulfed entity. But not this time. This time, the two cells formed an alliance, with the swallowed cell transferring many of its genes to its host, keeping only those involved in providing energy. The result – mitochondrion.

So, believe it or not, every animal really has two genomes – its standard DNA and the DNA inside of its mitochondria. However, the standard DNA inside the nucleus comes from a mashing of both parents’ DNA. The genetic material inside the mitochondria comes only from the mother.

Thus, by comparing the mtDNA from either twin, IU Anthropology professors Della Collins Cook and Frederika A. Kaestle could discover if they shared the same maternal lineage.

Not only were the two infants not twins, and therefore not possibly conjoined, but the two were not even maternal siblings, test results found. One infant belonged to Haplogroup C, an mtDNA lineage believed to have arisen geographically between the Caspian Sea and Lake Baikal about 60,000 years ago, and the other belonged to Haplogroup A, which is thought to have come from Asia between 30,000 and 50,000 years ago.

As to why the infants would have been buried together, then, when they weren’t related remains a mystery.